Abstract Myocardial infarction (MI) is caused by lack of adequate blood flow to the heart and results in terminal loss of cardiomyocytes (CM). MI and its related complications are a leading cause of death worldwide. The search for regenerative therapy that can repair or replace lost CM remains a daunting challenge. The foremost issue is the difficulty in procuring viable replacement cells. Heterologous sources are unattractive due to the potential for immunorejection. Harvesting progenitor cells from a patient's damaged heart is invasive and poses severe risks. Reprogramming somatic cells from other autologous sources can result in very low yields due to genetic or epigenetic barriers. A series of recent studies, however, has revealed that masseter muscle derived progenitor (MMP) cells share a common origin and have overlapping gene expression patterns with heart muscle. MMP can be isolated from highly accessible masseter muscles without mandible motor dysfunction and our preliminary data has shown that MMP yield the highest rate of CM differentiation as compared with limb muscle progenitors. Importantly, MMP can give rise to functional CM phenotypes including ventricular, atrial, and pacemaker CM under defined conditions. Therefore, MMP represent an ideal therapeutic candidate to maximize cardiogenic differentiation efficiency after cell transplantation in order to repopulate an infarcted heart region with a supply of autologous CM. At the same time, MMP avoid the common pitfalls associated with immunorejection, tumor formation, and reversion to an alternative epigenetic precursor. Aim 1 consists of in vitro studies to isolate and characterize MMP (including developmental origin, surface markers, and proliferation potential) in order to gain the desired CM population using novel sorting approaches. Aim 2 is designed to determine the mechanism by which microRNAs and transcription factor networks mediate the lineage commitments of MMP and the underlying cardiac potential of MMP as regulated by miR-128. Finally, Aim 3 focuses on the effects of implantation of MMP-derived cell sheets on the cardiac functions in mouse and porcine MI models. Experiments will examine the in vivo cell fate of MMP and determine any beneficial effects that result from cell engraftment and functional integration under ischemic conditions. These studies will provide new insights in both basic heart developmental biology and cell-based regenerative medicine. This approach holds great promise for the emerging field of personalized medicine and strongly supports the possibility that autologous MMP harvested from human masseter muscles and expanded in vitro will serve as a major source of CM that will be highly effective for treatment of patients after MI.